Effect of a Plant-derived Natural Compound, Flavin7, on the ERK Signaling Pathway in Immortalized Mouse Proximal Tubule Cells
Abstract. Background: Since MAP kinases represent an important pathway of transducing external stimuli to internal signals in cells, determining their possible role in cancer cells may offer a promising way for the treatment and prognosis of malignant diseases. Our previous experiments have shown that a flavonoid-rich solution, Flavin7, was able to diminish kidney tumor growth in vivo. Materials and Methods: Effects of Flavin7 on the MAPK signaling pathway were determined in immortalized mouse proximal tubule cells by determining cell viability, flow cytometric analysis, luciferase assays and Western blots. Results: At a nontoxic dose, Flavin7 markedly reduced phosphorylation of ERK and inhibited activity of its downstream targets such as Elk1 and CREB via inhibition of the ERK-kinase MEK1. Conclusion: Because of its ability to temporarily inhibit kidney tumor growth and activation of the MEK1/ERK pathway in vitro, further in vivo investigations may determine the potential role of Flavin7 in the treatment of malignancies. The mitogen-activated protein kinase (MAPK) cascade is a major signaling system by which cells transduce extracellular signals into intracellular responses. Many steps of this cascade are conserved, and homologs have been discovered in different species (1). The first three mammalian MAP kinases, ERK1, ERK2 and ERK3 were cloned in the early 1990s, facilitating the development of reagents for their study. It has become clear that ERK1 and ERK2 are among the protein kinases most commonly activated in signal transduction pathways. They have particularly been linked to cell proliferation, but have important roles in many other events (2-4). In mammalian cells, ERK1 and ERK2, often referred to as p44 and p42 MAP kinases, are the archetypal members of the MAPK family. Therefore, determining the possible role of MAPKs in cancer cells may offer a promising way for treatment and prognosis of cancerous diseases. According to recent results, activation of the ERK pathway is a frequent event in tumorigenesis. ERK has been implicated in cell initiation, tumor promotion and progression, invasion, metastasis, and regulation of apoptosis and angiogenesis, events that are essential for successful completion of developing a metastatic tumor (reviewed in 5). On the other hand, ERK activation is not unambiguously an advantage or a disadvantage for patients with cancerous diseases, since it has been shown to trigger cell proliferation and survival in normal cells, as well as in tumor cells. Flavonoids, found in great quantity in fruit extracts, are secondary metabolites of superior plants exhibiting antitumor effects. They are known to exert antioxidant and antiproliferative effects on tumor cells (6). Recent studies have speculated that the classical antioxidant activity of flavonoids is unlikely to be the sole explanation for their cellular effects. This hypothesis is based on several lines of reasoning: i) flavonoids are extensively metabolized in vivo, thus, their redox potentials are significantly altered (7), and ii) the concentrations of flavonoids and their metabolites accumulated in vivo are lower than those of small of antioxidant nutrients (8). Investigations have indicated that flavonoids may selectively interact with the MAPK signaling pathway due to their ability to inhibit tyrosine kinase activity (9, 10). A natural compound, Flavin7 (F7), composed of the extracts from seven different fruits, was investigated in our kidney tumor animal model (11). Ne/De tumor cells were transplanted underneath the renal capsule of 6- to 8-weekold Fisher344 rats and animals were treated with human 871 Correspondence to: Edit Nádasi, MD, Ph.D., Department of Public Health and Preventive Medicine, University of Pécs, 7624 Pécs, Szigeti u. 12, Hungary. Tel: +36 72 536394, Fax: +36 72 536395, e-mail: email@example.com Key Words: Flavin7 extract, plant-derived natural compounds, mitogen-activated protein kinase pathway, anticarcinogenic effect. in vivo 21: 871-876 (2007) Effect of a Plant-derived Natural Compound, Flavin7, on the ERK Signaling Pathway in Immortalized Mouse Proximal Tubule Cells EDIT NÁDASI1,2, ISTVÁN EMBER2 and ISTVÁN ARANY1 1Department of Internal Medicine, University of Arkansas for Medical Sciences and Central Arkansas Veteran HealthCare System, Little Rock, AR 72205, U.S.A.; 2Department of Public Health and Preventive Medicine, University of Pécs, 7624 Pécs, Hungary 0258-851X/2007 $2.00+.40 dose-equivalent F7 solution according to the manufacturer’s instructions. After two weeks of treatment rats were sacrificed and tumor growth was determined. F7 significantly (p<0.05) reduced tumor growth in vivo. Accordingly, the aim of this study was to determine whether F7 influences the ERK signaling pathway in immortalized mouse renal proximal tubule cells.